Respiratory syncytial virus (RSV) can cause lower respiratory illness (LRI) in otherwise healthy children and developing an effective prophylactic vaccine to mitigate RSV disease in infants is a public and global health priority according to the World Health Organization. The overall goal of this project is to develop a safe and efficacious RSV vaccine for immunizing sero-negative 4 months to 2 years old infants, who are beyond the reach of protective RSV maternal antibodies, against RSV hospitalizations and medically attended LRI. Globally, there are 34 million new RSV infections annually with 3.4 million hospitalizations and 66,000 to 199,000 deaths. Although most people are infected in the first year of life, recurring RSV infections occur throughout life. It was estimated that in the US, 2 million children under the age of 5 required care for RSV infections annually and 78% are over the age of one year. In the US, medical costs for managing RSV disease in infants were estimated at $1.15 billion annually with additional lost income and caregiver costs estimated at $625 million annually. Vaccinating the birth cohort with a RSV vaccine that has 50% efficacy and protection for 12 months was estimated to reduce medical costs by $236 million and income and productivity losses by $134 million annually in the US. A live attenuated RSV vaccine mimics natural RSV infection. RSV epidemiology and natural history studies have demonstrated that natural infection results in lasting protection against severe RSV disease. The Moore lab recently published on a vaccine candidate that was uniquely engineered to replicate like wild-type virus in infected cells and yet is less able to spread because of the down-regulation of virulence genes. The attenuation phenotype is stable because it is conferred by hundreds of mutations in the RSV genome. The other distinguishing characteristic is a proprietary fusion protein that is enriched for the more immunogenic pre- fusion conformation. The combination of these properties resulted in high levels of attenuation in cotton rats with almost wtRSV level neutralizing antibodies and complete protection against wtRSV challenge making this a promising vaccine candidate for clinical development. In Phase 1 the objectives are to generate research stocks of two live attenuated RSV vaccine candidates called OE4 and DB1Quad and establish a pre-seed process that can be transferred to a cGMP facility for initiation of vaccine production. In Phase 2, we will evaluate attenuation of OE4 and DB1Quad in human airway epithelial cells ex vivo and in cotton rats in vivo and perform studies in cotton rats to verify that they do not cause RSV vaccine-enhanced disease. The outcome of these studies will allow Meissa complete IND-enabling pharmacology studies and to prioritize a lead candidate for clinical development.